The invention relates to methods and/or compositions for the prevention and/or treatment of PCV-2-caused myocarditis, and/or abortion and/or intrauterine infection, as well as pathologic sequelae including but not limited to post-weaning multisystemic wasting syndrome; and, to methods for preparing such compositions and kits for preparing such compositions or for performing such methods, inter alia.
Various documents are cited in this text. Citations in the text can be by way of a citation to a document in the reference list, e.g., by way of an author(s) and document year citation to a document listed in the reference list, or by full citation in the text to a document that may or may not also be listed in the reference list.
There is no admission that any of the various documents cited in this text are prior art as to the present invention. Any document having as an author or inventor person or persons named as an inventor herein is a document that is not by another as to the inventive entity herein. All documents cited in this text (xe2x80x9cherein cited documentsxe2x80x9d) and all documents cited or referenced in herein cited documents are hereby incorporated herein by reference.
Porcine circovirus-2 (PCV-2) was recently identified as an agent that has been consistently associated with post-weaning multisystemic wasting syndrome (PMWS) in swine populations in several parts of the world (Allan et al. 1998; Ellis et al., 1998). Isolates of PCV-2 obtained from infected pigs in several countries are virtually identical genetically, and are distinctly different from the PCV (CCL33, PCV-1) that was originally identified in the 1970""s as a noncytopathic contaminant of porcine kidney (PK/15) cell line (Meehan et al. 1998; Tischer et al. 1974). Pigs with naturally acquired or experimentally induced PCV-2 infections present with progressive weight loss, tachypnea, dyspnea, and jaundice (Allan et al. 1998; Allan et al. 1999; Ellis et al. 1998; Ellis et al. 1999). Gross pathologic findings that have been directly associated with PCV-2 antigen include, lymphadenopathy, interstitial pneumonia, hepatitis and nephritis (Allan et al. 1998; Allan et al. 1999; Ellis et al. 1998; Ellis et al. 1999). PCV-2 has not heretofore been directly linked to abortion or lesions in fetal pigs. Thus, heretofore, it has not been proposed to address the issue of PCV-2-caused myocarditis, and/or abortion and/or intrauterine infection.
It has surprisingly been found that PCV-2 is a causative agent of myocarditis, abortion and intrauterine infection, as well as post-weaning multisystemic wasting syndrome.
By definition, a PCV-2 immunogen is intended to encompass live attenuated or inactivated PCV-2, or subunit(s) from PCV-2 obtained by in vitro expression or by extraction, or fragment(s) comprising at least one epitope of interest which can be obtained by chemical synthesis or by in vitro recombinant expression, as well as recombinant vector(s) comprising and expressing in vivo sequence(s) or fragment(s) or epitope(s) of PCV-2 genome as herein disclosed or as in documents cited or referenced herein.
A similar definition applies for an immunogen of another porcine pathogen as disclosed herein.
Thus, an object of the invention can be to provide methods and/or compositions for the prevention and/or treatment of PCV-2-caused myocarditis, and/or abortion and/or intrauterine infection, as well as post-weaning multisystemic wasting syndrome and/or pathologic sequelae including but not limited to post-weaning multisystemic wasting syndrome; and, methods for formulating such compositions and uses of a PCV-2 immunogen (which compositions can also include a porcine parvovirus (PPV) immunogen, wherein when recombinant vector expression is used, the vector can co-express both the PPV and the PCV-2 immunogens, inter alia) for formulating such compositions.
Another object of the invention is the isolation and characterisation of new PCV-2 strains identified 1103 (1103/1 P.2) and 1121 (1121/1 P.1), and their uses to produce immunogens, as well as antigens and antibodies for diagnostics, in relation with PCV-2-caused myocarditis, and/or abortion and/or intrauterine infection, as well as post-weaning multisystemic wasting syndrome and/or pathologic sequelae associated therewith.
The invention provides also for inoculation of female pigs (e.g., sows, gilts) with a composition comprising a (at least one) PCV-2 immunogen (which composition can also include an immunogen from porcine parvovirus) prior to breeding; and/or prior to serving, and/or during gestation (or pregnancy); and/or prior to the perinatal period or farrowing; and/or repeatedly over a lifetime , to prevent myocarditis and/or abortion and/or intrauterine infection associated with PCV-2, as well as post-weaning multisystemic wasting syndrome and other pathologic sequelae associated with PCV-2; or, to elicit an immunogenic or protective response against PCV-2 and thereby prevent post-weaning multisystemic wasting syndrome and/or myocarditis and/or abortion and/or intrauterine infection associated with porcine circovirus-2 and/or other pathologic sequelae associated with PCV-2.
Advantageously, at least one inoculation is done before serving. It is also advantageously followed by an inoculation to be performed during gestation, e.g., at about mid-gestation (at about 6-8 weeks of gestation) and/or at the end of gestation (at about 11-13 weeks of gestation). Thus, an advantageous regimen is an inoculation before serving and a booster inoculation during gestation. Thereafter, there can be reinoculation before each serving and/or during gestation at about mid-gestation (at about 6-8 weeks of gestation) and/or at the end of gestation (at about 11-13 weeks of gestation). Preferably, reinoculation can be during gestation only.
In another preferred embodiment, piglets, such as piglets from vaccinated females (e.g., inoculated as herein discussed), are inoculated within the first weeks of life, e.g., inoculation at one and/or two and/or three and/or four and/or five weeks of life. More preferably, piglets are first inoculated within the first week of life or within the third week of life (e.g., at the time of weaning). Even more advantageous, such piglets are then boosted two (2) to four (4) weeks later (after being first inoculated). Thus, both offspring, as well as female pig (e.g., sow, gilt) can be administered compositions of the invention and/or can be the subject of performance of methods of the invention.
Thus, the invention also comprehends immunogenic or vaccine compositions for preventing or treating myocarditis and/or abortion and/or intrauterine infection associated with porcine circovirus-2, as well as post-weaning multisystemic wasting syndrome and other pathologic sequelae associated with PCV-2. An immunogenic (or immunological) composition elicits an immunological responsexe2x80x94local or systemic. A vaccine composition elicits a local or systemic protective response. The terms xe2x80x9cimmunological compositionxe2x80x9d and xe2x80x9cimmunogenic compositionxe2x80x9d include a xe2x80x9cvaccine compositionxe2x80x9d (as the two former terms can be protective compositions). The composition can comprise a PCV-2 immunogen (which composition can. also include a PPV immunogen).
And, the invention further comprehends uses of a PCV-2 immunogen (which composition can also include a PPV immunogen) to formulate an immunogenic or vaccine composition for preventing or treating myocarditis and/or abortion and/or intrauterine infection associated with porcine circovirus-2, as well as post-weaning multisystemic wasting syndrome and other pathologic sequelae associated with PCV-2.
Further still, the invention comprehends an immunogenic or vaccine composition for the prevention and/or treatment of PCV-2-caused myocarditis, and/or abortion and/or intrauterine infection and/or post-weaning multisystemic wasting syndrome comprising a pharmaceutically or veterinarily acceptable carrier and/or vehicle and/or excipient and/or adjuvant, and a PCV-2 immunogen
The composition can additionally include at least one immunogen from at least one additional pig pathogen, e.g.: Porcine Reproductive and Respiratory Syndrome (PRRS), Mycoplasma hyopneumoniae, Actinobacillus pleuropneumoniae, E. coli, Bordetella bronchiseptica, Pasteurella multocida, Erysipelothrix rhusiopathiae, Pseudorabies, Hog cholera, Swine Influenza, and Porcine Parvovirus (PPV). Thus, vector-based compositions can include at least one immunogen from at least one additional pig pathogen, such as a vector expressing a sequence from this pathogen, wherein the vector can also be the vector expressing the PCV-2 immunogen. The vector expressing a PCV-2 sequence can comprise a PCV-2 sequence or fragment thereof as herein disclosed or as in documents cited or referenced herein; and the invention comprehends such nucleic acid molecules, vectors containing them, compositions comprising such nucleic acid molecules or vector expression products from such nucleic acid molecules, compositions comprising such expression products, probes or primers for such nucleic acid molecules, and methods for making and using any or all of the foregoing.
The vector can comprise a DNA vector plasmid, a bacteria such as an E. coli, a virus such as baculovirus, a herpesvirus including pig herpes viruses, including Aujeszky""s disease virus, an adenovirus including a porcine adenovirus, a poxvirus, including a vaccinia virus, an avipox virus, a canarypox virus, a racoonpox and a swinepox virus, and the like. The vector-based compositions can comprise a vector that contains and expresses an ORF selected from the group consisting of ORFs 1 to 13, such as an ORF selected from ORFs 4, 7, 10 and 13; preferably ORFs 4 and/or 13, of a PCV-2, advantageously of any one of the PCV-2 strains identified herein. And, the immunogen in compositions (either PCV-2 and/or from another pig pathogen) can be recombinantly produced. The word plasmid is intended to include any DNA transcription unit in the form of a polynucleotide sequence comprising the PCV sequence to be expressed. Advantageously, the plasmid includes elements necessary for its expression; for instance, expression in vivo. The circular plasmid form, supercoiled or otherwise, is advantageous; and, the linear form is also included within the scope of the invention. The plasmid immunogenic or vaccine composition can be administered by way of a gene gun, intradermally via an needleless injector, subcutaneously or intramuscularly, or by mucosal route, or by any other means that allows for expression in vivo, and advantageously an immunogenic or protective response.
It is noted that the expression product generated by vectors or recombinants in this invention optionally can also be isolated and/or purified from infected or transfected cells; for instance, to prepare compositions for administration to pigs; however, in certain instances, it may be advantageous not to isolate and/or purify an expression product from a cell; for instance, when the cell or portions thereof enhance the immunogenic effect of the polypeptide. And, techniques for protein purification and/or isolation from this disclosure and documents cited herein, inter alia, and thus within the ambit of the skilled artisan, can be used, without undue experimentation, to purify and/or isolate recombinant or vector expression products and/or subunits of PCV-2 and/or other pig pathogens, in the practice of the invention, and such techniques, in general, can include: precipitation by taking advantage of the solubility of the protein of interest at varying salt concentrations, precipitation with organic solvents, polymers and other materials, affinity precipitation and selective denaturation; column chromatography, including high performance liquid chromatography (HPLC), ion-exchange, affinity, immunoaffinity or dye-ligand chromatography; immunoprecipitation, gel filtration, electrophoretic methods, ultrafiltration and isoelectric focusing, and their combinations, inter alia.
The invention further envisages methods for the prevention and/or treatment of porcine circovirus-2 (PCV-2)-caused myocarditis, and/or abortion and/or intrauterine infection and/or post-weaning multisystemic wasting syndrome and/or other pathologic sequelae associated with PCV-2 comprising inducing an immunogenic or protective response against PCV-2 in a pig comprising administering to the pig an aforementioned or herein disclosed composition.
Thus, the invention comprehends a method for the prevention and/or treatment of porcine circovirus-2 (PCV-2)-caused myocarditis, and/or abortion and/or intrauterine infection and/or post-weaning multisystemic wasting syndrome and/or other pathologic sequelae associated with PCV-2 comprising inducing an immunogenic or protective response against PCV-2 in a pig comprising administering to the pig a composition comprising a pharmaceutically or veterinarily acceptable carrier or excipient or vehicle, with preferably an adjuvant, and an active agent comprising a PCV-2 immunogen. The method can be for the prevention of PCV-2-caused mycarditis and/or abortion and/or intrauterine infection comprising administering a composition comprising a pharmaceutically or veterinarily acceptable carrier and a PCV-2 immunogen. The PCV-2 immunogen can be an attenuated live whole PCV-2 or inactivated PCV-2. The method can involve a composition that is a subunit immunogenic, or vaccine composition. The method can involve the composition additionally including at least one immunogen from at least one additional pig pathogen , including a vector expressing such an immunogen or epitope; e.g., the at least one additional pig pathogen can be selected from the group consisting of PRRS, Mycoplasma hyopneumoniae, Actinobacillus pleuropneumoniae, E. coli, Pseudorabies, Hog cholera, Bordetella bronchiseptica, Pasteurella multocida, Erysipelothrix rhusiopathiae, Swine Influenza, and PPV and combinations thereof. The method can involve a vector that is a DNA vector plasmid, a bacteria such as an E. coli, a virus such as baculovirus, a herpesvirus including Aujeszky""s disease virus, an adenovirus including a porcine adenovirus, a poxvirus, including a vaccinia virus, an avipox virus, a canarypox virus, and a swinepox virus, and the like. The method can involve a vector-based composition additionally including at least one sequence, fragment or epitope from at least one additional pig pathogen, or a vector expressing such a sequence, fragment or epitope, wherein the vector can also be the vector expressing the PCV-2 sequence, fragment or epitope. The method can involve a vector that contains and expresses an ORF selected from the group consisting of ORFs 1 to 13, e.g., an ORF selectred from ORFs 4, 7, 10 and 13; preferably ORFs 4 and/or 13. The method can also involve an immunogen based composition wherein one or more of the immunogen(s) is recombinantly produced. In this method, females and/or piglets are inoculated as described above.
In another embodiment, the invention involves a method for preparing any of the aforementioned or herein disclosed compositions comprising admixing the pharmaceutically or veterinarily acceptable carrier and the PCV-2 immunogen. The method can further include transfecting or infecting a cell or host with a recombinant vector that contains DNA encoding a PCV-2 immunogen and expresses that immunogen; and optionally purifying and/or isolating the immunogen from the cell. Similarly the method can include isolating and/or purifying a PCV-2 immunogen from PCV-2, or isolating PCV-2 from a sample.
The invention also provides a kit for preparing any of the aforementioned or herein disclosed compositions or for performing any of the aforementioned or herein disclosed methods comprising in a first container the pharmaceutically or veterinarily acceptable carrier or vehicle or excipient and in a second container the active agent comprising the PCV-2 immunogen, wherein the first and second containers are optionally packaged together, and the kit optionally includes instructions for admixture of ingredients of the composition and/or administration of the composition.
In yet another embodiment, the invention provides for administering any of the aforementioned or herein disclosed compositions to male and/or female pigs; to prevent transmission of PCV-2 and prevent or treat or control myocarditis and/or abortion and/or intrauterine infection associated with porcine circovirus-2, as well as post-weaning multisystemic wasting syndrome and other pathologic sequelae associated with PCV-2. Administration is preferably done as described above.
The term xe2x80x9ccomprisingxe2x80x9d in this disclosure can mean xe2x80x9cincludingxe2x80x9d or can have the meaning commonly given to the term xe2x80x9ccomprisingxe2x80x9d in U.S. Patent Law.
Other aspects of the invention are described in or are obvious from (and within the ambit of the invention) the following disclosure.